Lawsonia intracellularis (also called Lawsonia) is the causative agent of proliferative enteropathy (also called enteritis or ileitis) in many animals, in particular pigs, and presents a clinical sign and pathological syndrome with mucosal hyperplasia of immature crypt epithelial cells, primarily in the terminal ileum. Other sites of the intestines that can be affected include the jejunum, caecum and colon. Weanling and young adult pigs are principally affected with typical clinical manifestation of rapid weight loss and dehydration. Natural clinical disease in pigs occurs worldwide. The disease is consistently associated with the presence of intracellular curved bacteria, presently known as Lawsonia intracellularis. In general, vaccination against Lawsonia intracellularis has shown to be an economically efficient measure to treat an infection with Lawsonia intracellularis. This way Ileitis can be controlled which allows a better exploitation of the genetic growth potential of the pig. At present there is only one vaccine on the market to protect against Lawsonia intracellularis, viz. Enterisol® Ileitis marketed by Boehringer Ingelheim. This vaccine is a live vaccine for oral administration. Other vaccines have also been described but are not commercially available. In any case, for commercial production of substantive volumes of a vaccine to treat an infection with Lawsonia intracellularis it would be desirable to have an efficient and economically attractive cultivation method for these intracellular bacteria.
From EP 843 818 a method is known to grow Lawsonia intracellularis bacteria in McCoy cells adhered to a substrate (such as a flask bottom or micro-carriers). Indeed, McCoy cells have proven to be adequate host cells to cultivate lawsonia intracellularis bacteria. It is also known from this patent that for passage of a culture to actually grow the Lawsonia intracelluaris bacteria (i.e. to increase the net number of viable bacteria), one needs to incubate fresh uninfected McCoy cells at low oxygen, that is below 18%, whereafter these fresh cells are inoculated with already infected McCoy cells. Passage to fresh McCoy cells is believed to be necessary since the infected McCoy host cells are ultimately killed by the intracellular Lawsonia bacteria. Indeed, it has been described that a Lawsonia infection spreads via host cells that lyse and therewith release the intracellular bacteria such that they become available for infection of other cells. A disadvantage of the known method is that at passage, the fresh cells have to be infected. This is a critical step, which takes a relatively long period of time (typically from 6 days up to two weeks but possibly even longer than that) and has a relatively high degree of uncertainty towards success. Due to these typical issues, additional infection steps therefore increase the costs of Lawsonia intracellularis cultivation significantly. Moreover, with every additional infection step Lawsonia intracellularis may acquire a higher level of attenuation. This makes the known cultivation technique less attractive for the production of bacteria for use in vaccines. Indeed, these disadvantages have been recognized in the art and therefore in WO 2009/088878 infected McCoy cells are not passed to fresh (uninfected) McCoy cells for growing Lawsonia intracellularis bacteria. Rather, infected McCoy cells are grown until full infection and then harvested. This way, the repeated infection step when passing the infected McCoy cells can be avoided.